This invention relates to a method for purification and/or concentration of antihemophilic factor (AHF). More particularly, this invention relates to a method for improving the final AHF product recovered from cryoprecipitated blood plasma using cold precipitation.
The process of blood coagulation is a complicated biological activity and involves the interaction of several substances found in normal whole blood. It is known that certain factors associated with the blood coagulation mechanism are absent or seriously deficient in certain individuals. For example, classical hemophilia (hemophilia A) is a disease caused by a deficiency of AHF (Factor VIII). In individuals suffering from the congenital hemophilia known as hemophilia B, the blood is deficient in plasma thromboplastin component (PTC, Factor IX). Several other factors which are important in the coagulation mechanism are Factors II, VII and X.
In the past, treatment of hemophiliacs consisted of transfusing the patient with whole blood or blood plasma. Better medical practice dictates that, whenever possible, the patient be administered only those blood components in which he is deficient. Due to the universal shortage of blood, it is also advantageous to fractionate blood into its various components, whereby individual components can be used for patient treatment as required.
Various methods of fractionating blood and blood plasma into its separate components or concentrates thereof are known. The work on development of the alcohol fractionation method is particularly noteworthy. With specific reference to the production of AHF, recent U.S. Pat. Nos. 4,383,989; 4,386,068; 4,387,092; 4,445,301 and 4,404,131 illustrate improved methods of obtaining a highly purified concentrate of AHF.
Unfortunately, there are a number of problems associated with the isolation of AHF. AHF is a glycoprotein that closely associates itself with other proteins in blood plasma, such as fibrinogen and fibronectin. Fibrinogen and fibronectin are considered contaminants in the final AHF solution. Therefore, it is necessary to precipitate out of the plasma as much fibrinogen and fibronectin as possible while maintaining a high specific activity of AHF in the final product. It is also desirable to obtain as high a yield of AHF as possible from the initial blood plasma due to the rarity and expense of whole blood. It is further advantageous to have a final AHF product with high solubility, because a low solubility of AHF requires the administration of large amounts of fluid to a patient. Large quantities of fluid are both a strain on the heart and circulation of the patient and make the hemophilia patient dependent on the continual substitution of the missing blood factors or infusions, as well as requiring the assistance of physicians for administration. High fibrinogen and fibronectin contect in the final product results in lower solubility than is desired.
Various methods are known in the art for isolating AHF, but these methods are often extremely expensive to perform and do not provide an optimum combination in the final AHF product of high solubility, low fibrinogen and fibronectin content, high specific activity of AHF and high yield of AHF from the initial blood plasma. What is needed therefore, is a method isolating AHF which meets all of these criteria and is not prohibitively expensive. The present invention satisfies these needs and provides other related advantages.